Temperature-modulated Diversity of TRPV4 Channel Gating

Abstract

The TRPV4 calcium-permeable channel was cloned from mouse kidney M-1 cells, and the effect of temperature modulation on channel gating/activation by physical and chemical signals was evaluated. A TRPV4 cDNA construct with a C-terminal V5 epitope was stably transfected into human embryonic kidney (HEK) 293 and Chinese hamster ovary cells resulting in high levels of expression at the plasma membrane. Channel activation was assessed from changes in calcium influx (fura-2 fluorescence measurements) or whole cell currents (patch clamp analysis). At room temperature (22-24°C), exposure of TRPV4-transfected cells to hypotonic medium (225 mOsm/liter) or a non-protein kinase C (PKC)-activating phorbol ester derivative, 4␣-phorbol 12,13-decanoate (100 nM), induces modest channel activation, whereas phorbol 12-myristate 13-acetate (100 nM), a PKC-activating phorbol ester, and shear stress (3-20 dyne/cm 2 ) had minimal or no effect on channel activation. In contrast, at elevated temperatures (37°C) the channel was rapidly activated by all stimuli. Inhibition of PKC by calphostin C (50 nM) or staurosporine (500 nM) abolished phorbol 12-myristate 13-acetate-induced activation of the channel without affecting the response to other stimuli. Ruthenium red (1 M) effectively blocked the channel activity by all stimuli. It is concluded that temperature is a critical modulator of TRPV4 channel gating, leading to activation of the channel by a diverse range of microenvironmental chemical and physical signals utilizing a least two transduction pathways, one PKC-dependent and one PKC-independent. The convergence of multiple signals and transduction pathways on the same channel indicate that the channel functions as a molecular integrator of microenvironmental chemical and physical signals.The TRP calcium-permeable cation channels are a rapidly growing superfamily of channels expressed in a broad range of both excitable and nonexcitable cells. The channels, in general, are not voltage-activated but appear to be activated by a broad range of ligands and intracellular mediators (1-7). The superfamily has been divided into three main subfamilies based on structural and functional similarities: TPRVC, the canonical TRP channels first identified in Drosophila; TRPV, the vanilloid subfamily named after its first member, the vanilloid receptor (or capsaicin receptor; renamed TRPV1); and TRPM, the melatonin subfamily named after its first member, melatonin (see Refs. 1 and 8 for nomenclature). The TRPV subfamily has demonstrated a notable broad sensitivity to chemical and physical stimuli, particularly for TRPV1, which is activated by various noxious mediators, including heat, acid, and capsaicin (the hot ingredient of peppers), and by endogenous lipid mediators, such as diacylglycerol and anandamide (2, 5, 9). Other members of the TRPV subfamily have been shown, so far, to have a narrower range of sensitivities: heat and insulinlike growth factor-I for TRPV2 (10, 11); heat for TRPV3 (12)(13)(14); and vitamin D and related ligands for TRPV5 (4)...

Warm Temperatures Activate TRPV4 in Mouse 308 Keratinocytes

“…We previously demonstrated that expression of TRPV4 in keratinocytes of the rat plantar epidermis was not confined to the plasma membrane but was diffusely distributed throughout the cytoplasm, excluding the nucleus (10). Other investigators have demonstrated that a pool of recombinant TRPV4 is expressed in an apparently intracellular compartment in transfected fibroblasts (24). TRPV3 appears to exhibit a similar pattern in skin sections (3), a result we have confirmed.…”

Angiotensin II Increases Activity of the Epithelial Na+ Channel (ENaC) in Distal Nephron Additively to Aldosterone

“…[Ca 2ϩ ] i Measurements-Intracellular calcium levels were measured in cells of the split-opened ASDNs using Fura-2 fluorescence ratiometric imaging as described previously (32,33). Split-opened ASDNs were loaded with Fura-2 by incubation with 2 M Fura-2/AM in a bath solution for 40 min at room temperature.…”

Activation of the TRPV4 Ion Channel Is Enhanced by Phosphorylation

“…TRPV4 is a member of the transient receptor potential vanilloid subfamily of TRP 2 channels, and like other members of this subfamily, it is a polymodal receptor activated by a wide variety of stimuli. TRPV4 is strongly expressed in kidney and is activated by hypotonicity, which has led to the suggestion that TRPV4 is an osmosensor important in regulating body fluid levels (2,(5)(6)(7)(8)(9). However, TRPV4 is also activated by innocuous heat with a threshold of Ͼ27°C (6,10,11), by the phorbol ester 4␣-phorbol 12,13-didecanoate (12,13), by low pH (14), by endocannabinoids and arachidonic acid metabolites (15,16), by the active compound, bisandrographolide A, of Andrographis paniculata, a Chinese herbal plant (17), and by nitric oxide (18).…”